In this project we determine the antigenic relationships based on VP4 and VP7 neutralization specificities of various rotavirus strains derived from humans and animals. The elucidation of the neutralization specificities of rotaviruses is important in order to achieve a more comprehensive understanding of rotavirus epidemiology and for formulation of an effective strategy for vaccination. Of 5 globally important VP7 (G) serotypes (G1-4 and 9) of rotaviruses, G9 continues to attract considerable attention. That is because the G9 viruses have a unique natural history and evolution in human communities. Serotype G9 rotavirus was isolated from a child with diarrhea first in the US in 1983 and subsequently in Japan in 1985. Curiously, soon after their detection, G9 rotaviruses were not detected for about a decade in both countries and then reemerged in both countries in the mid-1990s. Unexpectedly, however, such reemerged G9 strains were distinct genetically and molecularly from those isolated in the 1980s. Thus, the origin of the reemerged G9 viruses remains an enigma. Sequence analysis has demonstrated that the G9 rotavirus VP7 gene belongs to one of at least three phylogenetic lineages: lineage 1 (strains isolated in the 1980s in the US and Japan), lineage 2 (strains first isolated in 1986 and exclusively in India thus far) and lineage 3 (strains that emerged or reemerged in the mid-1990s). Currently, lineage 3 G9 viruses are the most frequently detected G9 strains globally. Previously, we characterized a porcine G9 rotavirus and reported that the VP7 of the porcine G9 virus was closely related serotypically and molecularly to contemporary human lineage 3 virus VP7s and suggested that the porcine G9 VP7 and human lineage 3 G9 virus VP7s may have a common ancestor or perhaps the porcine G9 virus VP7 itself may be a progenitor of contemporary lineage 3 human virus VP7s. During rotavirus strain surveillance of archival diarrheal stool samples collected at Childrens Hospital in Washington DC from 1974 to 1991, we detected 3 G9 viruses in samples collected in 1980. We successfully grew two of the three 1980 G9 viruses in cell cultures and then characterized them biologically and molecularly. These viruses were shown to be closely related serotypically to lineage 3 G9 viruses and distantly related to lineages 1 and 2 G9 viruses. However, the VP7 gene of these G9 viruses was found to have (i) a closest phylogenetic relationship to lineage 2 G9 virus VP7 gene, (ii) a close phylogenetic relationship to lineage 1 G9 virus VP7 genes, and (iii) a distant phylogenetic relationship to contemporary lineage 3 G9 virus VP7 genes. Moreover, codon usage bias analyses of the VP7 gene of a total of 26 G9 viruses (obtained from GenBank database) confirmed such findings. These two 1980 viruses, which are the earliest G9 viruses isolated in humans thus far, together with lineage 2 G9 virus may represent ancestral viruses from which lineage 1 G9 viruses evolved.